Phosphotidylinositol-3-Kinase Inhibitor (PIK75) Enhanced Nnanosystems For Anticancer Drug Delivery

Abstract

Development of new anticancer agents is pursued with keen interest by health researchers for cancer therapy. However several such compounds which are discovered have poor water solubility which limits their preclinical and clinical development. Using formulation strategies to improve solubility, dissolution, delivery and efficacy of these agents is an effective strategy to investigate the application of these compounds in preclinical studies. PIK75 is a potent and specific PI3K p110α inhibitor with poor water solubility which has limited its preclinical investigation. The aim of this study was to design and evaluate formulation systems of PIK75. Two nano formulation systems were investigated for the delivery of PIK75 namely nanosuspension and nanoemulsion system. The targeted delivery of nanosuspension and nanoemulsions system was also investigated for effective delivery of PIK75 to cancer cells. A pro-apoptotic molecule C6-ceramide was also co-delivered to augment therapeutic efficacy. Preliminary formulation studies with PIK75 were conducted by preparing PIK75 nanosuspension formulations using high pressure homogenisation technique. The physicochemical properties of the nanosuspension were characterized and pharmacokinetic and tissue distribution parameters were investigated. Poloxamer-188 and soybean lecithin were used to stabilize the nanoparticles and a 32 factorial design was established to optimize the homogenising parameters. PIK75 nanosuspension with an average particle size of 160-180 nm, entrapment efficiency of 97-98% and surface charge above -30mV were prepared. The nanosuspension showed an 11-fold improvement in saturation solubility and stability recovery was greater than 90% for 6 h in the nanosuspension system and in human plasma. In vivo, PIK75-NS showed a similar plasma pharmacokinetic profile to PIK75 suspension, but tissue distribution studies indicated lower PIK75 levels in the liver and kidney post PIK75-NS administration. In the second part of this investigation in order to improve drug delivery through targeting, PIK75 surface functionalized nanosystems were developed and evaluated. EGFR and FR functionalized nanoemulsions incorporating PIK75 and C6-ceramide were characterized for particle size, surface charge, entrapment efficiency and morphology. Fluorescence and quantitative uptake studies were conducted in SKOV-3 cells to determine intracellular distribution. Cell viability was assessed using MTT assay while mechanism of cytotoxicity was evaluated using capsase-3/7, TUNEL and hROS assay. Cytotoxicity assay showed a 57% decrease in IC50 value of PIK75 following treatment with EGFR targeted nanoemulsion and a 40% decrease following treatment with FR targeted nanoemulsion. Combination therapy with PIK75 and ceramide enhanced the cytotoxicity of PIK75 compared to therapy with individual formulations. The increase in cytotoxicity was attributed to increase in cellular apoptosis and hROS activity. PIK75 nanosuspension was also surface functionalized with folic acid to prepare targeted nanosuspension. In vitro dissolution testing showed improved dissolution rate and increase in saturation solubility (161 fold for PIK75 NS and 113 fold for PIK75-FA NS compared to solubility in PBS). In vitro studies in SKOV-3 cells indicated improved uptake and 62% decrease in IC50 value of PIK75 following treatment with targeted nanosuspension compared to non-targeted nanosuspension. These results illustrated the opportunity to formulate PIK75 as a targeted delivery system for effective uptake and cytotoxicity in ovarian cancer.